Application of a Microsphere-Based Array for Rapid Identification of Acinetobacter spp. with Distinct Antimicrobial Susceptibilities

Abstract

Acinetobacter spp. have emerged as important nosocomial and multidrug-resistant pathogens in the last decade. A. calcoaceticus, A. baumannii, Acinetobacter genospecies 3, and Acinetobacter genospecies 13TU are genetically closely related and are referred to as the A. calcoaceticus-A. baumannii complex (ACB complex). Distinct Acinetobacter spp. may be associated with differences in antimicrobial susceptibility, so it is important to identify Acinetobacter spp. at the species level. We developed a microsphere-based array that combines an allele-specific primer extension assay and microsphere hybridization for the identification of Acinetobacter spp. This assay can discriminate the 13 different Acinetobacter spp. in less than 8.5 h, and it has high specificity without causing cross-reactivity with 14 other common nosocomial bacterial species. The sensitivity of this assay was 100 A. baumannii cells per ml of blood, and it could discriminate multiple species in various mixture ratios. The developed assay could differentiate clinical Acinetobacter spp. isolates with a 90% identification rate. The antimicrobial susceptibility test showed that A. baumannii isolates were resistant to most antimicrobial agents other than imipenem, while the genospecies 3 and 13TU isolates were more susceptible to most antimicrobial agents, especially ciprofloxacin and ampicillinsulbactam. These results supported the idea that this assay possibly could be applied to clinical samples and provide accurate species identification, which might be helpful for clinicians when they are treating infections caused by Acinetobacter spp.

The Ecology, Biology and Pathogenesis of Acinetobacter spp.: An Overview

“…A recent diagnostic method which was reported to have high specificity and can discriminate between Acinetobacter species is the microsphere-based array technique that combines an allelespecific primer extension assay and microsphere hybridization (73). The method was reported to be so efficient that 13 different species of Acinetobacter were discriminated in less than 9 h with 90% accuracy and precision (73). The use of DNA-DNA hybridization and sequence analysis (19,49) is considered the gold standard, but the method is laborintensive and impractical in most clinical laboratories.…”

“…from environmental samples Vanbroekhoven et al have been employed (112). A recent diagnostic method which was reported to have high specificity and can discriminate between Acinetobacter species is the microsphere-based array technique that combines an allelespecific primer extension assay and microsphere hybridization (73). The method was reported to be so efficient that 13 different species of Acinetobacter were discriminated in less than 9 h with 90% accuracy and precision (73).…”

Safety Assessment of Transgenic Organisms, Volume 4

“…A PCR-and hybridization-based method using microspheres and flow-cytometric-based detection could be used to discriminate amongst 13 different Acinetobacter spp. and detect as few as 100 A. baumannii cells per mL blood (Lin et al, 2008). A highthroughput method based on PCR-ligase reaction and capillary electrophoresis was described which could detect A. baumannii in blood (Pingle et al, 2007).…”

“…Other insecticidal genes from Bt have been introduced (especially cry1Ac and cry2Ab, and more recently vip3A), which encode particular proteins selectively toxic to various Lepidopteran pests. An insecticidal gene AaHIT from the scorpion Androctonus australis is also being explored in transgenic cotton against some lepidopterans (Wu et al, 2008).…”

“…Other insecticidal genes from Bt have been introduced (especially cry1Ac and cry2Ab, and more recently vip3A), which encode particular proteins selectively toxic to various Lepidopteran pests. An insecticidal gene AaHIT from the scorpion Androctonus australis is also being explored in transgenic cotton against some lepidopterans (Wu et al, 2008).Cotton lines have been genetically engineered to tolerate the herbicides bromoxynil, glufosinate ammonium, glyphosate or sulfonylurea. Transgenic cottons in commercial production include plants having stacked transgenes, for example a Bt (Cry1Ac + Cry2Ab) + glyphosate-tolerant cotton.…”